Molecular Cloning of an Inulin Fructotransferase (Depolymerizing) Gene fromArthrobactersp. H65–7 and Its Expression inEscherichia coli

1 January 1997

journal article
Published by Taylor & Francis in Bioscience, Biotechnology, and Biochemistry

Vol. 61 (1) , 87-92
https://doi.org/10.1271/bbb.61.87

Abstract

The gene encoding an extracellular inulin fructotransferase (depolymerizing) (inulase II) (EC 2.4.1.93), designated ift gene, was cloned from the genomic DNA of Arthrobacter sp. H65-7, and expressed in Escherichia coli for the first time. Sequence analysis showed a single open reading frame consisting of 1314 base pairs that encoded a signal peptide of 32 amino acids and a mature protein of 405 amino acids. The primary structure showed a homology of 49.8% with that of the inulin fructotransferase (DFA I-producing) (EC 2.4.1.200) from Arthrobacter globiformis S14-3. E. coli cells carrying the ift gene produced the active enzyme under control of the lac promoter. The expression of the ift gene was improved by a plasmid, pIFT-B, in which the ift gene was immediately downstream from the lac promoter. An E. coli transformant carrying pIFT-B expressed twice as much activity of inulase II as that of the original strain, Arthrobacter sp. H65-7. Most of the enzyme activity existed within the cells.

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